The functional analysis of cell polarity protein aPKCI. in glomerular podocytes

我们最初通过足细胞特异性缺失aPKClambda建立了肾小球疾病的小鼠模型。这些小鼠的血液、尿液和肾脏的临床和组织病理学检查证实，它们出现了裂隙隔膜功能障碍，并伴有蛋白尿、肾小球硬化和肾功能衰竭。这些结果表明，我们的小鼠可以作为分析肾小球疾病的有用模型。我们扩展了这些结果来揭示肾小球疾病的分子机制，得到了以下结果。在不同阶段对模型小鼠的电镜分析揭示了裂隙横膈膜缺陷的逐步发展。虽然裂隙横膈膜最初形成，但随着小鼠的生长，裂隙横膈膜逐渐脱位、紊乱、消失。肾小球基底膜未见明显变化。为了阐明aPKC在调节肾小球功能中的作用，我们研究了aPKC与狭缝膈结构蛋白nephrin和podocin之间的功能相互作用。我们发现aPKC的特异性抑制剂显著干扰了离体大鼠肾小球中nephrin和podocin的分布。而aPKC抑制剂对nephrin-podocin复合物的形成无明显影响。这些数据表明，aPKC是结构蛋白在狭缝隔膜中合理分布所必需的。这也提示aPKC的功能紊乱可能参与肾小球疾病的发生。通过以上观察，我们发现aPKC调节特定脂质的限制来确定基底外侧结构域，aPKC结合蛋白PAR3在顶端结构域的建立中起关键作用。我们公布并报告了这些结果，如第11节所示。

We originally established a mouse model of glomerular disease by podocyte-specific deletion of aPKClambda. Clinical and histopathological examinations of blood, urine, and kidneys of these mice confirmed that they develop dysfunctions of the slit diaphragm with proteinuria, glomerulosclerosis, and renal failure. These results indicate that our mice can serve as a useful model to analyze glomerular diseases. We extended these results to reveal molecular mechanisms of glomerular diseases and obtained the following results.1. The electron microscopic analyses of our model mice in various stages revealed a step-wise progression of defects in the slit diaphragms. Although the slit diaphragms were formed at first, they were dislocated, disorganized, and lost along with the growth of mice. The glomerular basement membranes were not significantly affected.2. To clarify the function of aPKC in the regulation of glomerular functions, we examined the functional interactions between aPKC and the structural proteins in slit diaphragms: nephrin and podocin. We fund that a specific inhibitor for aPKC significantly disturbed the distribution of nephrin and podocin in the isolated rat glomeruli. However, the formation of nephrin-podocin complex was not significantly affected by aPKC inhibitor.These data indicate that aPKC is required for the proper distribution of the structural proteins in slit diaphragms. It is also suggested that the functional disturbance in aPKC can be involved in the development of glomerular diseases.Along with above observations, we revealed that aPKC regulates the restriction of a specific lipid to determine the basolateral domains and that aPKC-binding protein PAR3 plays a critical role in the establishment of apical domains.We published and reported these results as indicated in the section 11.